Expression of the Voltage- and Ca 21 -Dependent BK Potassium Channel Subunits BKb1 and BKb4 in Rodent Astrocytes KATHARINA N. SEIDEL, 1 CHRISTIAN DERST, 1 MIKHAIL SALZMANN, 1 MARKUS H € OLTJE, 1 JOSEF PRILLER, 2,3 REN  E MARKGRAF, 4 STEFAN H. HEINEMANN, 4 HEIKE HEILMANN, 1 SERGUEI N. SKATCHKOV, 5,6 MISTY J. EATON, 5,6 R € UDIGER W. VEH, 1 AND HARALD PR € USS 2,7 * 1 Institute for Integrative Neuroanatomy, Charit  e University Medicine Berlin, 10115 Berlin, Germany 2 Department of Neuropsychiatry and Laboratory of Molecular Psychiatry, Charit  e University Medicine Berlin, 10117 Berlin, Germany 3 Berlin-Brandenburg Center for Regenerative Therapies, Augustenburger Platz 1, 13353 Berlin, Germany 4 Department of Biophysics, Center for Molecular Biomedicine, Friedrich Schiller University of Jena, University Hospital Jena, 07745 Jena, Germany 5 Department of Biochemistry, Universidad Central del Caribe, School of Medicine, Bayam on, Puerto Rico 6 Department of Physiology, Universidad Central del Caribe, School of Medicine, Bayam on, Puerto Rico 7 Department of Neurology and Experimental Neurology, Charit  e University Medicine Berlin, 10117 Berlin, Germany KEY WORDS large-conductance Ca 21 -activated potassium channels; regulatory BKb subunit; astrocytes; Bergmann glia; marginal glia; neurovascular coupling; membrane potential ABSTRACT Large-conductance Ca 21 -activated (BK) potassium channels are centrally involved in neurovascular coupling, immunity, and neural transmission. The ability to be synergistically acti- vated by membrane depolarization, different ligands and intracellular Ca 21 links intracellular signaling and mem- brane excitability. The diverse physiological functions of BK channels crucially depend on regulatory b subunits. Although first studies characterized the neuronal distribu- tion of BKb subunits in the rodent brain, it is largely unknown which b subunit proteins are expressed in astro- cytes and thus mediate these regulatory effects. We there- fore analyzed the expression of BKb subunits in rat and mouse brain and glial cell cultures. A monospecific polyclo- nal antibody against the BKb4 channel subunit was raised, affinity-purified and extensively characterized. BKb4 and to a lesser degree BKb1 transcripts and protein were detected in several astrocytic populations and cultured cells. Particu- larly strong BKb4 immunostaining was detected in astro- cytic progenitors derived from the subventricular zone. The overlapping expression of BKa and BKb4 in astrocytes implies a functional relationship and suggests that BKb4 is an important accessory b subunit for astrocytic BK chan- nels. In addition, BKb4 might exert effects independent of the a subunit as functional heterologous co-expression of Nav1.6 and BKb4 resulted in reduced Nav1.6 sodium cur- rents. Thus, BKb4 expression in astrocytes likely partici- pates in regulating astrocytic voltage gradients and maintaining K 1 homeostasis, hence enabling astrocytes to fulfill their complex regulatory influence on proper brain function. Ó 2011 Wiley-Liss, Inc. INTRODUCTION Large-conductance Ca 21 -activated potassium channels (BK, MaxiK, Slo1) are widely distributed ion channels assembled by four pore-forming a subunits, which are centrally involved in many physiological functions such as blood flow, immunity, and neural transmission (Lu et al., 2006). The ability to be synergistically acti- vated by membrane depolarization, intracellular Ca 21 and other ligands makes them unique and links intra- cellular signaling pathways to membrane excitability (Rothberg, 2004). They are key molecules in neurovascu- lar coupling releasing K 1 from astrocytic endfeet into the perivascular space after increase in astrocytic intra- cellular Ca 21 (Filosa et al., 2006). This finding revealed that BK channels centrally participate in the complex regulation along the neurovas- cular unit consisting of neurons, glial cells, and vascula- ture. It further corroborates that astrocytes are not just of structural importance in the brain, but rather actively help to control accurate brain function including the neu- ronal activity-dependent regulation of cerebral blood flow. Beyond astrocytic endfeet, the glial expression of BK channels has not been emphasized, although several studies analyzed the neuronal distribution in the rodent brain (Gu et al., 2007; Knaus et al., 1996). BK currents have been detected electrophysiologically in rat astro- cytes in vitro (Gebremedhin et al., 2003) and the expres- sion of BKa protein was seen in murine astrocyte primary cultures (Ou et al., 2009). The diverse physiological functions of BK channels largely depend on regulatory b subunits (Brenner et al., 2005; Torres et al., 2007). At present, four BKb subunits have been cloned in mammals, which regulate the Ca 21 and voltage sensitivity of BK channels (Torres et al., 2007). b subunits mediate the effect of different signal Grant sponsor: DFG (grant number HE2993/5); BMBF/BCRT; DFG/NeuroCure. *Correspondence to: Harald Pr€ uss, Department of Neurology and Experimental Neurology, Charit e University Medicine Berlin, Charit eplatz 1, 10117 Berlin, Germany. E-mail: harald.pruess@charite.de Received 19 July 2010; Accepted 20 January 2011 DOI 10.1002/glia.21160 Publishedonline 24 March 2011 in Wiley Online Library (wileyonlinelibrary.com). GLIA 59:893–902 (2011) V V Ó 2011 Wiley-Liss, Inc.